, 2011). The KO slice cultures suffered less neuronal cell death than WT cultures subjected to the same duration of OGD, consistent with the idea that the function of these molecules in the brain—in neurons or possibly other resident glial or microglia—normally contributes to damage after stroke. Future experiments involving cell-type-specific knockout mice will help dissect the relative contribution of peripheral immune cells, neurons, and brain glia to damage and impaired recovery. KbDb KO mice, offspring of breeding pairs on a C57BL/6 background, were generously provided Selleck Autophagy Compound Library by H. Ploegh (Vugmeyster et al., 1998 and Ziskin et al., 2007). C57BL/6 (i.e., KbDb WT) controls were purchased
(Charles River). PirB KO and PirB WT controls were previously generated in C.J.S.’s laboratory (Syken et al., 2006). Mice were maintained in a pathogen-free environment. All experiments using animals were performed blind to genotype and in accordance with a protocol approved by the Stanford University Animal Care and Use Committee and in keeping with the National Institutes of Health’s Guide for the Care and Use of Laboratory
Animals. Transient ischemia was induced in male mice (postnatal days 60–90) by using the suture occlusion technique, as previously described (Han et al., 2009), with slight modifications. This age was chosen because it is beyond the developmental critical periods, but the animals are still relatively young adults. See Supplemental
Experimental Procedures isothipendyl for complete details of experimental procedures. We thank N. Sotelo and P. Kemper for help with laboratory logistics and mice breeding, Dr. H. Lee and Selleck VE822 Y. Kim for additional troubleshooting for qPCR reactions, and S. Cheng for genotyping assistance. The PirB mutant mice were generated by Dr. J. Syken in the Shatz laboratory. Thanks also to Dr. H. Ploegh, MIT, for KbDb KO mice. This work was supported by NIH grants MH07166 and EY02858, the Mathers Charitible Foundation, and the Ellison Foundation to C.J.S., a National Defense Science and Engineering Graduate Fellowship and National Science Foundation Graduate Research Fellowship to J.D.A., and NIH grants RO1 GM49831 and NS 053898 to R.G.G. “
“Activity-evoked and spontaneous vesicle exo-/endocytosis support the two fundamental modes of neurotransmission at synapses. Within our current understanding, the arrival of an action potential at the synapse triggers an increase in the cytosolic Ca2+ concentration, which leads to evoked fusion of vesicles docked at the active zone within a few milliseconds (Sudhof, 2004). Such activity-evoked vesicle fusion supports most of neurotransmitter release and is an essential component of synaptic information transmission. However, it is also known that vesicle fusion and uptake occur even in the absence of activity, albeit at a very low rate of one to two vesicles per minute (Geppert et al., 1994 and Murthy and Stevens, 1999).